Pharmaceutical Formulation for Histone Deacetylase Inhibitors

ABSTRACT

A pharmaceutical composition, comprising a therapeutically effective amount of an active pharmaceutical ingredient (API) compound represented by the following structural formula 
     
       
         
         
             
             
         
       
         
         
           
             at least one acidifying agent; and a vehicle base comprising at least one pharmaceutically acceptable non-aqueous solvent. Values and preferred values of the variables in structural formula (I) are defined herein.

RELATED APPLICATIONS

This application is a divisional of Ser. No. 16/275,395, filed Feb. 24,2019, which is a division of Ser. No. 15/637,979, now U.S. Pat. No.10,258,595, which is a divisional of Ser. No. 14/983,761, filed Dec. 30,2015, now abandoned, which is a continuation of Ser. No. 13/878,994, nowU.S. Pat. No. 9,255,066, which is a U.S. National Stage ofPCT/US2011/056148, filed Oct. 13, 2011, which claims the benefit of Ser.No. 61/392,855, filed on Oct. 13, 2010; the entire contents of which arehereby incorporated by reference.

BACKGROUND OF THE INVENTION

Post-translational modification of proteins through acetylation anddeacetylation of lysine residues has a critical role in regulatingcellular functions, making histone deacetylase an attractive biologicaltarget, particularly for the treatment of cancer. For example, WO2007/095584 describes small molecule inhibitors or histone deacetylasehaving an esterase sensitive ester linkage. The presence of theesterase-sensitive linker provides an inhibitor which can achieve highlocal concentrations and reduced systemic toxicity.

Some small molecules that include labile covalent bonds can be unstablein aqueous solvents. For example, the presence of an esterase-sensitivelinker may present difficulties with respect to preparation of asuitable formulation. As such, a need exists for stable pharmaceuticalcompositions of histone deacetylase inhibitors when the inhibitor haslabile covalent bonds, such as those described in WO 2007/095584.

SUMMARY OF THE INVENTION

The present invention relates to pharmaceutical composition comprising ahistone deacetylase inhibitor, methods of using such a pharmaceuticalcomposition and kits suitable for preparing such a pharmaceuticalcomposition.

In one embodiment, the present invention is a pharmaceutical compositioncomprising a therapeutically effective amount of an activepharmaceutical ingredient (API) compound represented by the followingstructural formula

at least one acidifying agent; and a vehicle base comprising at leastone pharmaceutically acceptable non-aqueous solvent. In structuralformula (I), n is an integer from 0 to 15, R₁ and R₂ are eachindependently hydrogen, halogen, an aliphatic group, a heteroaliphaticgroup, an aryl, a heteroaryl; —OR_(A); —C(O)R_(A); —C(O)N(R_(A))₂;—CO₂R_(A); —CN; —SCN; —SR_(A); —SOR_(A); —SO₂R_(A); —NO₂; —N(R_(A))₂;—NHR_(A); —NR_(A)C(O)R_(A); or —C(R_(A))₃; and wherein R_(A) for eachoccurrence is, independently, a hydrogen, an aliphatic group, aheteroaliphatic group, an acyl moiety, an aryl moiety, a heteroarylmoiety, alkoxy; aryloxy; alkylthio, arylthio, amino, alkylamino,dialkylamino, heteroaryloxy, or heteroarylthio moiety.

In another embodiment, the present invention is a pharmaceuticalcomposition. The composition comprises at least 0.01% by weight of anactive pharmaceutical ingredient (API) compound represented by thefollowing structural formula

at least one acidifying agent; and at least one pharmaceuticallyacceptable non-aqueous solvent. Preferably, n is 6, R₂ is hydrogen andR₁ is —C(O)OR_(A), R_(A) is a C₁-C₁₂ alkyl; the at least onepharmaceutically acceptable non-aqueous solvent is a mixture of ethanoland propylene glycol; the at least one acidifying agent is selected fromone or more of citric acid and phosphoric acid and the measured pH ofthe pharmaceutical composition is from about 3 to about 5 In aparticular aspect of this embodiment, the pharmaceutical compositionfurther optionally includes one or more of the following: at least onehumectant selected from glycerin and hexylene glycol; at least oneemollient selected from diisopropyl adipate and oleyl alcohol; at leastone permeation enhancer selected from ethanol, propylene glycol, oroleyl alcohol; a hydroxypropylcellulose gelling agent; and a butylatedhydroxytoluene as an antioxidant.

In another embodiment, the present invention is a method of treating adisorder in a subject in need thereof. The method comprises cutaneouslyadministering to the subject a therapeutically effective amount of apharmaceutical composition, wherein the disorder is selected from aproliferative disorder, an immune disorder and a skin disorder. Thepharmaceutical composition useful for practicing the method of thepresent invention is described above.

In another embodiment, the present invention is a kit. The kit comprisesa first container comprising an active pharmaceutical ingredient (API)compound represented by the following structural formula

and

a second container comprising a vehicle base comprising at least onepharmaceutically acceptable non-aqueous solvent. Values and preferredvalues of the variables in structural formula (I) are defined hereinbelow.

Pharmaceutical formulations of the present invention unexpectedlycombine effective API delivery with extended shelf life at roomtemperature. A vehicle base comprising an acidified non-aqueous solventcontributes to the shelf life extension by retarding the degradation ofAPI, for example by cleavage (e.g. by hydrolysis or solvolysis) of thelabile covalent bonds of the API molecule.

BRIEF DESCRIPTION OF THE DRAWINGS

The foregoing will be apparent from the following more particulardescription of example embodiments of the invention, as illustrated inthe accompanying drawings in which like reference characters refer tothe same parts throughout the different views. The drawings are notnecessarily to scale, emphasis instead being placed upon illustratingembodiments of the present invention.

The FIGURE is a block diagram of one embodiment of the manufacturingprocess that can be employed to manufacture a pharmaceutical compositionof the present invention.

DETAILED DESCRIPTION OF THE INVENTION

Histone deacetylase inhibitors such as those described in WO2007/095584, incorporated herein by reference in its entirety, are smallmolecules that include labile covalent bonds that can be unstable inaqueous solvents. It has now been surprisingly discovered that stabilityand shelf-life of pharmaceutical formulations comprising the histonedeacetylase inhibitors of WO 2007/095584 can be significantly enhancedby employing non-aqueous solvents that have been acidified.

In particular, some of the API compounds described in WO 2007/095584include a labile bond (e.g. an ester bond), which hydrolyzes at roomtemperature. The mechanism of this hydrolysis may be eitherbase-catalyzed or acid-catalyzed, depending on pH level. Traditionalmethods of retarding hydrolysis include either adjusting acidity (pH) ofan aqueous solvent or substituting an aqueous solvent with a non-aqueoussolvent. However, available experimental data indicates that neither oneof these methods alone provides the desired API stability. Surprisingly,the combination of at least one non-aqueous solvent and at least oneacidifying agent provides the desired API stability. Discovery of thesuitability of this combination is unexpected. More particularly,because the rate of hydrolysis of a labile bond in different solvents asa function of acidity is unpredictable, the discovery of the existenceof a combination of a solvent and an acidifying agent that provide for asufficient stability of an API is also unpredictable and, therefore,unexpected.

Definitions of Terms

The term “aliphatic”, as used herein, means non-aromatic group thatconsists solely of carbon and hydrogen and may optionally contain one ormore units of unsaturation, e.g., double and/or triple bonds. Analiphatic group may be straight chained, branched or cyclic.

The term “heteroaliphatic,” as used herein, means an aliphatic group inwhich one or more carbon atoms is replaced with a heteroatom, e.g., byO, N, S, Si, P or the like.

The term “alkyl”, as used herein, unless otherwise indicated, meansstraight or branched saturated monovalent hydrocarbon radicals offormula C_(n)H_(2n+1). In some embodiments, n is from 1 to 18. In otherembodiments, n is from 1 to 12. Preferably, n is from 1 to 6. In someembodiments, n is 1-1000, for example, n is 1-100. Alkyl can optionallybe substituted with —OH, —SH, halogen, amino, cyano, nitro, a C₁-C₁₂alkyl, C₁-C₁₂ haloalkyl, C₁-C₁₂ alkoxy, C₁-C₁₂ haloalkoxy, C₁-C₁₂ alkylsulfanyl, C₂-C₁₂ alkenyl, C₂-C₁₂ alkynyl group, an aryl or a heteroaryl.In addition, an alkyl can be substituted with ═O, ═S, ═N-alkyl. Furtherexamples of suitable substituents on an alkyl group include —R′OR, whereeach R and each R′ is independently an alkyl, an aryl or a heteroarylgroup. The term alkyl can also refer to cycloalkyl.

As used herein, an “alkenyl group,” alone or as a part of a largermoiety (e.g., cycloalkene oxide), is preferably a straight chained orbranched aliphatic group having one or more double bonds with 2 to about12 carbon atoms, e.g., ethenyl, 1-propenyl, 1-butenyl, 2-butenyl,2-methyl-1-propenyl, pentenyl, hexenyl, heptenyl or octenyl, or acycloaliphatic group having one or more double bonds with 3 to about 12carbon atoms. Exemplary substituents of an alkenyl group are describedabove with respect to alkyl.

As used herein, an “alkynyl” group, alone or as a part of a largermoiety, is preferably a straight chained or branched aliphatic grouphaving one or more triple bonds with 2 to about 12 carbon atoms, e.g.,ethynyl, 1-propynyl, 1-butynyl, 3-methyl-1-butynyl,3,3-dimethyl-1-butynyl, pentynyl, hexynyl, heptynyl or octynyl, or acycloaliphatic group having one or more triple bonds with 3 to about 12carbon atoms. Exemplary substituents of an alkynyl group are describedabove with respect to alkyl.

The term “cycloalkyl”, as used herein, means saturated cyclichydrocarbons, i.e. compounds where all ring atoms are carbons. In someembodiments, a cycloalkyl comprises from 3 to 18 carbons. Preferably, acycloalkyl comprises from 3 to 6 carbons. Examples of cycloalkylinclude, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl,cyclohexyl, and cycloheptyl. Exemplary substituents of a cycloalkyl aredescribed above with respect to alkyl.

The term “haloalkyl”, as used herein, includes an alkyl substituted withone or more F, Cl Br, or I, wherein alkyl is defined above. Exemplarysubstituents of a haloalkyl are described above with respect to alkyl.

The terms “alkoxy,” as used herein, means an “alkyl-O—” group, whereinalkyl is defined above. Examples of alkoxy group include methoxy orethoxy groups. Exemplary substituents of an alkoxy are described abovewith respect to alkyl.

The terms “alkylthio,” as used herein, means an “alkyl-S—” group,wherein alkyl is defined above. Examples of alkylthio group includeCH₃—S— or CH₃—CH₂—S— groups. Exemplary substituents of an alkylthiogroup are described above with respect to alkyl.

The term “aryl,” as used herein, refers to a carbocyclic aromatic group.Preferably, an aryl comprises from 6 to 18 carbons. Examples of arylgroups include, but are not limited to phenyl and naphthyl. Examples ofaryl groups include optionally substituted groups such as phenyl,biphenyl, naphthyl, phenanthryl, anthracenyl, pyrenyl, fluoranthyl orfluorenyl. An aryl can be optionally substituted on a substitutablecarbon atom. Examples of suitable substituents on an aryl includehalogen, hydroxyl, cyano, nitro, C1-C12 alkyl, C2-C12 alkenyl or C2-C12alkynyl, C1-C12 haloalkyl, C1-C12 alkoxy or haloalkoxy group, an aryloxygroup, an arylamino group, an aryl, a heteroaryl group. In addition, anaryl can be substituted with ═O, ═S, ═N-alkyl. Further examples ofsuitable substituents on an aryl group include —R′OR, where each R andeach R′ is independently an alkyl, an aryl or a heteroaryl group.

In some embodiments, a C6-C18 aryl is selected from the group consistingof phenyl, indenyl, naphthyl, azulenyl, heptalenyl, biphenyl, indacenyl,acenaphthylenyl, fluorenyl, phenalenyl, phenanthrenyl, anthracenyl,cyclopentacyclooctenyl or benzocyclooctenyl. In some embodiments, aC6-C14 aryl selected from the group consisting of phenyl, naphthalene,anthracene, 1H-phenalene, tetracene, and pentacene.

The term “aryloxy,” as used herein, means an “aryl-O—” group, whereinaryl is defined above. Examples of an aryloxy group include phenoxy ornaphthoxy groups. Suitable substituents on an aryloxy group are asdefined above with respect to an aryl group.

The term “arylthio,” as used herein, means an “aryl-S—” group, whereinaryl is defined above. Examples of an aryloxy group include phenylthioor naphthylthio groups. Suitable substituents on an arylthio group areas defined above with respect to an aryl group.

The term “heteroaryl,” as used herein, refers to aromatic groupscontaining one or more heteroatoms (O, S, or N). A heteroaryl group canbe monocyclic or polycyclic, e.g. a monocyclic heteroaryl ring fused toone or more carbocyclic aromatic groups or other monocyclic heteroarylgroups. The heteroaryl groups of this invention can also include ringsystems substituted with one or more oxo moieties. Examples ofheteroaryl groups include, but are not limited to, pyridinyl,pyridazinyl, imidazolyl, pyrimidinyl, pyrazolyl, triazolyl, pyrazinyl,quinolyl, isoquinolyl, tetrazolyl, furyl, thienyl, isoxazolyl,thiazolyl, oxazolyl, isothiazolyl, pyrrolyl, quinolinyl, isoquinolinyl,indolyl, benzimidazolyl, benzofuranyl, cinnolinyl, indazolyl,indolizinyl, phthalazinyl, pyridazinyl, triazinyl, isoindolyl, purinyl,oxadiazolyl, thiazolyl, thiadiazolyl, furazanyl, benzofurazanyl,benzothiophenyl, benzotriazolyl, benzothiazolyl, benzoxazolyl,quinazolinyl, quinoxalinyl, naphthyridinyl, dihydroquinolyl,tetrahydroquinolyl, dihydroisoquinolyl, tetrahydroisoquinolyl,benzofuryl, furopyridinyl, pyrolopyrimidinyl, and azaindolyl.

In other embodiments, a 5-14-membered heteroaryl group selected from thegroup consisting of pyridyl, 1-oxo-pyridyl, furanyl, benzo[1,3]dioxolyl,benzo[1,4]dioxinyl, thienyl, pyrrolyl, oxazolyl, imidazolyl, thiazolyl,a isoxazolyl, quinolinyl, pyrazolyl, isothiazolyl, pyridazinyl,pyrimidinyl, pyrazinyl, a triazinyl, triazolyl, thiadiazolyl,isoquinolinyl, indazolyl, benzoxazolyl, benzofuryl, indolizinyl,imidazopyridyl, tetrazolyl, benzimidazolyl, benzothiazolyl,benzothiadiazolyl, benzoxadiazolyl, indolyl, tetrahydroindolyl,azaindolyl, imidazopyridyl, quinazolinyl, purinyl,pyrrolo[2,3]pyrimidinyl, pyrazolo[3,4]pyrimidinyl,imidazo[1,2-a]pyridyl, and benzothienyl.

The foregoing heteroaryl groups may be C-attached or N-attached (wheresuch is possible). For instance, a group derived from pyrrole may bepyrrol-1-yl (N-attached) or pyrrol-3-yl (C-attached).

The term “(hetero)arylthio,” as used herein, means an “(hetero)aryl-S—”group, wherein aryl is defined above. Examples of an arylthio groupinclude phenylthio or naphthylthio groups.

Suitable substituents for heteroaryl are as defined above with respectto aryl group.

In some embodiments, suitable substituents for a substitutable carbonatom in an aryl, a heteroaryl or an aryl portion of an arylalkenylinclude halogen, hydroxyl, C1-C12 alkyl, C2-C12 alkenyl or C2-C12alkynyl group, C1-C12 alkoxy, aryloxy group, arylamino group and C1-C12haloalkyl.

In the context of the present invention, an amino group may be a primary(—NH₂), secondary (—NHR_(p)), or tertiary (—NR_(p)R_(q)), wherein R_(p)and R_(q) may be any of the alkyl, alkenyl, alkynyl, alkoxy, cycloalkyl,cycloalkoxy, aryl, heteroaryl, and a bicyclic carbocyclic group. A(di)alkylamino group is an instance of an amino group substituted withone or two alkyls.

A “trialkylamino” group is a group —N⁺(R_(t))₃, wherein R_(t) is analkyl, as defined above.

As used herein, the term “acyl” refers to alkanoyl, i.e. a nalkyl-C(O)—, where “alkyl” is defined above.

The term (hetero)arylamine, as used herein, means an “(hetero)aryl-NH—”,an “(hetero)aryl-N(alkyl)-”, or an “((hetero)aryl)₂-N—” groups, wherein(hetero)aryl and alkyl are defined above.

A “humectant,” as used herein, is an excipient that can increase thewater level in the upper layers of the skin. Examples of humectantsapproved for use in topical drug products by the FDA include, but arenot limited to, the following: hexylene glycol, propylene glycol,sorbitol, lactic acid, sodium lactate, methyl gluceth-10, methylgluceth-20, and polyethylene glycols.

An “emollients,” as used herein, is an excipient that can improve skinfeel by softening, lubricating, and refatting the skin. Emollients mayalso improve the barrier function of skin and reduce water evaporation.Examples of emollients approved for use in topical drug products by theUnited States Food and Drug Administration (FDA) include, but are notlimited to, the following: diisopropyl adipate, isopropyl myristate,isopropyl palmitate, cetearyl octonoate, isopropyl isostearate, myristyllactate, octyldodecanol, oleyl alcohol, mineral oil, petrolatum,vegetable/plaint oils (e.g., peanut, soybean, safflower, olive, almond,coconut), PPG-15 stearyl ether, PPG-26 oleate, PEG-4 dilaurate,lecithin, lanolin, lanolin alcohol, polyoxyl 75 lanolin, cholesterol,cetyl esters wax, cetostearyl alcohol, glyceryl monostearate, mediumchain triglyerides, dimethicone, and cyclomethicone.

An “acidifying agent,” as used herein, refers to a chemical compoundthat alone or in combination with other compounds can be used to acidifya vehicle base of a pharmaceutical composition comprising a non-aqueoussolvent. An acceptable acidifying agent for a topical formulation is anacid with a pKa of 9.5 or less, more preferably having a pKa of 7.0 orless, most preferably having a pKa of 5.0 or less. Examples ofacidifying agents approved for use in topical drug products by the FDAinclude, but are not limited to: acetic acid, dehydro acetic acid,ascorbic acid, benzoic acid, boric acid, citric acid, edetic acid,hydrochloric acid, isostearic acid, stearic acid, lactic acid, nitricacid, oleic acid, phosphoric acid, sorbic acid, sulfuric acid, tartaricacid, and undecylenic acid.

A “nonaqueous solvent,” as used herein, is a solvent other than water.Examples of nonaqueous solvents approved for use in topical drugproducts by the FDA include, but are not limited to: alcohol (ethanol),acetone, benzyl alcohol, diethylene glycol monoethyl ether, glycerin,hexylene glycol, isoproypl alcohol, polyethylene glycols,methoxypolyethylene glycols, diethyl sebacate, dimethyl isosorbide,propylene carbonate, and dimethyl sulfoxide. Many emollients that areliquid at room temperature can also be used as solvents. These include,but are not limited to: diisopropyl adipate, isopropyl myristate,vegetable/plant oils, mineral oil, and isopropyl palmitate.

An “antioxidant,” as used herein, is a substance that inhibits oxidationof chemical compounds. Examples of typical antioxidants include alphatocopherol (all isomers), beta tocopherol, delta tocopherol, gammatocopherol, tocopherols, ascorbic acid, ascorbyl palmitate, butylatedhydroxyanisol (BHA), butylated hydroxytoluene (BHT), fumaric acid, malicacid, methionine, propyl gallate, sodium ascorbate, sodiummetabisulfate, sodium thiosulfate, sodium bisulfate. The chemicalcompounds protected from oxidation by an antioxidant include activepharmaceutical ingredients (API) and excipients containing moietiessusceptible to oxidation.

A “gelling agent,” as used herein, is a compound that thickens (i.e.increases the viscosity of) a formulation. Examples arehydroxypropylcelluloses, carbomer, hydroxyethylcelluloses,carboxymethylcelluloses, xanthan gum, guar gum, chitosan, polyvinylalcohol, povidone, carrageenan, methyl cellulose, hydroxypropyl methylcellulose, fatty alcohols, cetyl alcohol, stearyl alcohol, cetostearylalcohol, and myristyl alcohol. In a particular embodiment, the gellingagent is a pharmaceutical grade hydroxypropylcellulose Klucel® MF(“Klucel MF PH”), available from Hercules Incorporated of WilmingtonDel., under the catalog number 494-9.

A “skin permeation enhancer,” as used herein, is a compound thatimproves absorption of a pharmaceutically active ingredient through acutaneous membrane, e.g. skin.

“Measured pH,” as used herein, is the acidity of an aliquot of apharmaceutical composition of the present invention as defined herein,measured by diluting the aliquot with water to about 10% by volume.

As used herein, the term “therapeutically effective amount” refers to anamount of the API which is sufficient to reduce or ameliorate theseverity, duration, progression, or onset of a disorder being treated,prevent the advancement of a disorder being treated, cause theregression of, prevent the recurrence, development, onset or progressionof a symptom associated with a disorder being treated, or enhance orimprove the prophylactic or therapeutic effect(s) of another therapy.The precise amount of compound administered to a subject will depend onthe type and severity of the disease or condition and on thecharacteristics of the subject, such as general health, age, sex, bodyweight and tolerance to drugs. It will also depend on the degree,severity and type of the disorder being treated. The skilled artisanwill be able to determine appropriate dosages depending on these andother factors. When co-administered with other agents, e.g., whenco-administered with an anti-proliferative agent, a “therapeuticallyeffective amount” of the second agent will depend on the type of drugused. Suitable dosages are known for approved agents and can be adjustedby the skilled artisan according to the condition of the subject, thetype of condition(s) being treated and the amount of a compound of theinvention being used. In cases where no amount is expressly noted, aneffective amount should be assumed.

As used herein, the terms “treat”, “treatment” and “treating” refer tothe reduction or amelioration of the progression, severity and/orduration of a disorder being treated, or the amelioration of one or moresymptoms (preferably, one or more discernible symptoms) of a disorderbeing treated resulting from the administration of one or morepharmaceutical compositions of the present invention. In specificembodiments, the terms “treat”, “treatment” and “treating” refer to theamelioration of at least one measurable physical parameter beingtreated. For example, for a proliferative disorder, such parameters caninclude growth of a tumor, not necessarily discernible by the patient.In other embodiments the terms “treat”, “treatment” and “treating” referto the inhibition of the progression of a disorder being treated, eitherphysically by, e.g., stabilization of a discernible symptom,physiologically by, e.g., stabilization of a physical parameter, orboth. In the embodiments in which the disorder being treated is aproliferative disorder, the terms “treat”, “treatment” and “treating”can refer to the reduction or stabilization of cancerous cell count.

An “ointment,” as used herein, is a semisolid dosage form, usuallycontaining less than 20% water and volatiles and more than 50%hydrocarbons, waxes, or polyols as the vehicle. This dosage form isgenerally for external application to the skin or mucous membranes.

“Semisolid,” as used herein refers to the aggregate state of the matterthat is not pourable; it does not flow or conform to its container atroom temperature; it does not flow at low shear stress and generallyexhibits plastic flow behavior.

A “gel,” as used herein, is a semisolid dosage form that contains agelling agent to thicken to a solution or fine particle dispersion.Thickening the formulation aids in application to a specific site on thebody. A gel may contain suspended particles.

“Fine particle dispersion,” as used herein, is a system in which fineparticles of (i.e. typically less than 10-50 μm) are distributeduniformly throughout a liquid.

A “foam,” as used herein, is a dosage form containing gas bubblesdispersed in a liquid that contains less than 50% water, and in aparticular embodiment, no water.

Pharmaceutical Compositions

In one embodiment, the present invention is a pharmaceuticalcomposition, comprising a therapeutically effective amount of an activepharmaceutical ingredient (API) compound represented by the followingstructural formula

at least one acidifying agent and a vehicle base comprising at least onepharmaceutically acceptable non-aqueous solvent. In structural formula(I), n is an integer from 0 to 15, R₁ and R₂ are each independentlyhydrogen, halogen, an aliphatic group, a heteroaliphatic group, an aryl,a heteroaryl; —OR_(A); —C(O)R_(A); —C(O)N(R_(A))₂; —CO₂R_(A); —CN; —SCN;—SR_(A); —SOR_(A); —SO₂R_(A); —NO₂; —N(R_(A))₂; —NHR_(A);—NR_(A)C(O)R_(A); or —C(R_(A))₃, wherein R_(A) for each occurrence is,independently, a hydrogen, an aliphatic group, a heteroaliphatic group,an acyl moiety, an aryl moiety, a heteroaryl moiety, alkoxy; aryloxy;alkylthio, arylthio, amino, alkylamino, dialkylamino, heteroaryloxy, orheteroarylthio moiety.

In some embodiments, in structural formula (I), n is 5, 6 or 7,preferably, n is 6. Values and preferred values of the remainder of thevariable are as defined above with respect to structural formula (I).

In other embodiments, in structural formula (I), R₂ is hydrogen and R₁is selected from a halogen, —OR_(A), N(R_(A))₂, —NHR_(A), —C₁-C₆ alkyl,—C(O)R_(A), —C(O )OR_(A), —C(O)N(R_(A))₂, —C(O)NH₂, —CHO and—NHC(O)R_(A). Values and preferred values of the remainder of thevariable are as defined above with respect to structural formula (I).Preferably, R_(A) is hydrogen or a C₁-C₁₂ alkyl. Values and preferredvalues of the remainder of the variable are as defined above withrespect to structural formula (I).

In other embodiments, in structural formula (I), n is 6, R₂ is hydrogenand R₁ is —C(O)OR_(A), and R_(A) is a C₁-C₁₂ alkyl. Values and preferredvalues of the remainder of the variable are as defined above withrespect to structural formula (I).

In some embodiments, at least one acidifying agent is selected from thegroups consisting of acetic acid, dehydro acetic acid, ascorbic acid,benzoic acid, boric acid, citric acid, edetic acid, hydrochloric acid,isostearic acid, stearic acid, lactic acid, nitric acid, oleic acid,phosphoric acid, sorbic acid, sulfuric acid, tartaric acid, undecylenicacid, fumaric acid, malic acid, maleic acid, benzene sulfonic acid,cyclamic acid, diatrizoic acid, deoxycholic acid, gentisic acid,glucuronic acid, glutamic acid, and succinic acid. In a particularembodiment, at least one acidifying agent is selected from one or moreof citric acid, acetic acid, and phosphoric acid.

In some embodiments, semi-polar non-aqueous solvents are preferred(e.g., miscible with water such as ethanol, propylene glycol). In otherembodiments, at least one pharmaceutically acceptable non-aqueoussolvent is selected from the groups consisting of ethanol, acetone,benzyl alcohol, 2-(2-ethoxyethoxy)ethanol, diethylene glycol monoethylether, glycerin, propylene glycol, propylene carbonate, acetone,hexylene glycol, isopropyl alcohol, polyethylene glycols (PEGs),methoxypolyethylene glycols, diethyl sebacate, dimethyl isosorbide,propylene carbonate, dimethyl sulfoxide (DMSO), diisopropyl adipate,isopropyl myristate, vegetable oils, a mineral oil, and isopropylpalmitate. In a particular embodiment, at least one pharmaceuticallyacceptable non-aqueous solvent is selected from ethanol, benzyl alcohol,propylene glycol, 2-(2-ethoxyethoxy)ethanol, hexylene glycol anddiisopropyl adipate.

In some embodiments, a pharmaceutical composition of the presentinvention further includes at least one humectant. At least onehumectant is selected from the groups consisting of hexylene glycol,glycerin, propylene glycol, sorbitol, lactic acid, sodium lactate,mannitol, butylene glycol, panthenol, hyaluronic acid, urea, chitosan,polyols, methyl gluceth-10, methyl gluceth-20, and polyethylene glycols.In a particular embodiment, at least one humectant is selected fromglycerin and hexylene glycol.

In some embodiments, the pharmaceutical composition of the presentinvention further includes at least one emollient. In some embodiments,at least one emollient is selected from the groups consisting ofdiisopropyl adipate, isopropyl myristate, isopropyl palmitate, cetearyloctonoate, isopropyl isostearate, myristyl lactate, octyldodecanol,oleyl alcohol, a mineral oil, petrolatum, a vegetable oil, PPG-15stearyl ether, PEG-4 dilaurate, lecithin, lanolin, lanolin alcohol,polyoxyl 75 lanolin, cholesterol, cetyl esters wax, cetostearyl alcohol,glyceryl monostearate, triglycerides of capric and caprylic acids,dimethicone, and cyclomethicone. In a particular embodiment, at leastone emollient is selected from diisopropyl adipate and oleyl alcohol.

In particular embodiments, the pharmaceutical composition of the presentinvention includes at least one humectant and at least one emollient.Values and preferred values of humectants and emollients are listedabove. In a particular embodiment, at least one humectant is selectedfrom one or more of glycerin and hexylene glycol and wherein the atleast one emollient is selected from diisopropyl adipate and oleylalcohol.

In some embodiments, the pharmaceutical composition of the presentinvention further includes at least one skin permeation enhancer.Suitable skin permeation enhancers include, but are not limited to,dimethylsulfoxide (DMSO), decylmethylsulfoxide, N,N-dimethyl acetamide,N,N-dimethyl formamide, 2-pyrrolidone, 1-methyl-2-pyrrolidone,5-methyl-2-pyrrolidone, 1,5-methyl-2-pyrrolidone, 1-ethyl-2-pyrrolidone,2-pyrrolidone-5 carboxylic acid, propylene glycol, ethanol, isopropanol,oleic acid, laurocapram (Azone), limonene, cineole, diethyl-m-toluamide(DEET), sodium dodecylsulfate, trimethyl phosphine oxide,tetrahydrofurfuryl alcohol, glycerol monolaurate, methyl oleate,propylene glycol monolaurate, and oleyl alcohol. In a specific aspect,at least one permeation enhancer is selected from oleyl alcohol,propylene glycol and ethanol.

In some embodiments, the pharmaceutical composition of the presentinvention further includes at least one gelling agent. In particularembodiments, gelling agents are neutral polymers that thicken at low pH,such as cellulose-based polymers, for example hydroxypropylcellulose. Inparticular embodiments, at least one gelling agents is selected fromhydroxypropylcellulose, carbomer, hydroxyethylcellulose,carboxymethylcellulose, xanthan gum, guar gum, chitosan, polyvinylalcohol, povidone, carrageenan, methyl cellulose, hydroxypropyl methylcellulose, fatty alcohols, cetyl alcohol, stearyl alcohol, cetostearylalcohol, and myristyl alcohol. In a particular embodiment, at least onegelling agent is a hydroxypropylcellulose such as Klucel MF PH.

In some embodiments, the pharmaceutical composition of the presentinvention further includes at least one antioxidant. In a particularembodiment, the antioxidant is selected from the groups consisting ofalpha tocopherol, beta tocopherol, delta tocopherol, gamma tocopherol,tocopherols, ascorbic acid, ascorbyl palmitate, butylated hydroxyanisol(BHA), butylated hydroxytoluene (BHT), fumaric acid, malic acid,methionine, propyl gallate, sodium ascorbate, sodium metabisulfate,sodium thiosulfate, sodium bisulfate. In a particular embodiment, theantioxidant is the butylated hydroxytoluene (BHT).

In various embodiments, the pharmaceutical composition of the presentinvention comprises at least 0.01% by weight of the API. In someembodiments, the pharmaceutical composition of the present inventioncomprises from about 0.01% to about 15% by weight of the API. In otherembodiments, the pharmaceutical composition of the present inventioncomprises at least about 0.1% by weight of the API, alternatively—atleast 0.5% by weight of the API. For example, the pharmaceuticalcomposition of the present invention comprises, by weight, at leastabout 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%,0.1%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1.0%, 1.5%,2%, 2.5%, 3%, 3.5%, 4%, 4.5% or 5% of the API. In certain embodiments,the pharmaceutical composition of the present invention comprises atleast about 10% by weight of the API.

In various embodiments, the measured pH of the pharmaceuticalcomposition of the present invention is about 7. In other embodiments,the measured pH if the pharmaceutical composition of the presentinvention is from about 3 to about 6. In a particular embodiment, themeasured pH is from about 3 to about 5.

An example of a pharmaceutical composition of the present invention, isa pharmaceutical composition as defined herein above, comprising atleast 0.01% by weight of the API, wherein the vehicle base comprises amixture of ethanol and propylene glycol, and wherein the measured pH ofthe pharmaceutical composition is from about 3 to 5. In a particularembodiment, the at least one acidifying agent is selected from one ormore of citric acid and phosphoric acid. In some embodiments, thepharmaceutical composition further includes at least one humectantselected from one of more of glycerin and hexylene glycol. In otherembodiment, the pharmaceutical composition further includes at least oneemollient selected from one or more of diisopropyl adipate and oleylalcohol. In a particular embodiment, the pharmaceutical compositionfurther includes at least one permeation enhancer selected from one ormore of oleyl alcohol, propylene glycol and ethanol. In furtherembodiments, the pharmaceutical composition further includes ahydroxypropylcellulose gelling agent such as Klucel MF PH.

In another example, a pharmaceutical composition of the presentinvention comprises at least 0.01% by weight (e.g., 0.01%, 0.02%, 0.03%,0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, 0.1%, 0.1%, 0.2%, 0.3%, 0.4%,0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1.0%, 1.5%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%or 5%) of an active pharmaceutical ingredient (API) compound representedby the following structural formula

wherein n is 6, R₂ is hydrogen and R₁ is —C(O)OR_(A), R_(A) is a C₁-C₁₂alkyl. The pharmaceutical composition can further include at least oneacidifying agent, and at least one pharmaceutically acceptablenon-aqueous solvent, the at least one pharmaceutically acceptablenon-aqueous solvent is a mixture of ethanol and propylene glycol, the atleast one acidifying agent is selected from one or more of citric acidand phosphoric acid and the measured pH of the pharmaceuticalcomposition is from about 3 to about 5. The pharmaceutical compositioncan further include at least one humectant selected from one of more ofglycerin and hexylene glycol; at least one emollient selected from oneor more of diisopropyl adipate and oleyl alcohol; at least onepermeation enhancer selected from one or more of oleyl alcohol,propylene glycol and ethanol; a hydroxypropylcellulose gelling agentsuch as Klucel MF PH; and a butylated hydroxytoluene as an antioxidant.

In a particular embodiment, the pharmaceutical composition of thepresent invention comprises the API is a compound represented by thefollowing structural formula

In the above-described compositions of the present invention, the amountof a pharmaceutically acceptable non-aqueous solvent is the amountsuitable to bring the composition to the desirable volume and/or weight.For example, on the weight per weight basis (% w/w), the amount of oneor more solvents is from about 5% to about 99.9%. For example, theamount of one or more solvents can be 5%, 10%, 15%, 20%, 25%, 30%, 35%,40%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 99%. In otherembodiments, on the weight per weight basis (% w/w), the amount of oneor more humectants is, for example, from about 5% to about 50%, e.g. 5%,10%, 15%, 20%, 25%, 30%, 35%, 40%, 50%. In other embodiments, on theweight per weight basis (% w/w), the amount of one or more emollientsis, for example, from about 5% to about 50%, e.g. 5%, 10%, 15%, 20%,25%, 30%, 35%, 40%, 50%. In other embodiments, on the weight per weightbasis (% w/w), the amount of one or more antioxidants is, for example,from about 0.01% to about 5%, e.g. 0.01%, 0.05, 0.1%, 0.15%. 0.2%, 0.5%,1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, or 5.0%. In other embodiments,on the weight per weight basis (% w/w), the amount of one or moreacidifying agents is, for example, from about 0.01% to about 5.0%, e.g.0.01%, 0.05, 0.1%, 0.15%. 0.2%, 0.5%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%,3.5%, 4.0%, or 5.0%. In other embodiments, on the weight per weightbasis (% w/w), the amount of one or more gelling agents is, for example,from about 0.5% to about 5.0%, e.g. 0.5%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%,3.5%, 4.0%, or 5.0%.

Transdermal, topical, and mucosal dosage forms of the invention include,but are not limited to, ophthalmic and otic solutions, sprays, foams,aerosols, creams, lotions, ointments, gels, solutions, emulsions,suspensions, or other forms known to one of skill in the art. See, e.g.,Remington's Pharmaceutical Sciences (1980 & 1990) 16th and 18th eds.,Mack Publishing, Easton Pa. and Introduction to Pharmaceutical DosageForms (1985) 4th ed., Lea & Febiger, Philadelphia. Further, transdermaldosage forms include “reservoir type” or “matrix type” patches, whichcan be applied to the skin and worn for a specific period of time topermit the penetration of a desired amount of active ingredients.

Suitable excipients (e.g., carriers and diluents) and other materialsthat can be used to provide transdermal, topical, and mucosal dosageforms encompassed by this invention are well known to those skilled inthe pharmaceutical arts, and depend on the particular tissue to which agiven pharmaceutical composition or dosage form will be applied. See,e.g., Remington's Pharmaceutical Sciences (1980 & 1990) 16th and 18theds., Mack Publishing, Easton, Pa.

Depending on the specific tissue to be treated, additional componentsmay be used prior to, in conjunction with, or subsequent to treatmentwith active ingredients of the invention. For example, penetrationenhancers can be used to assist in delivering the active ingredients tothe tissue.

Different salts, hydrates or solvates of the active ingredients can beused to further adjust the properties of the resulting composition.

In a particular embodiment, pharmaceutical compositions of the presentinvention can be formulated as non-aqueous ointments, non-aqueous gelsor non-aqueous foams.

Methods of Treatment

In some embodiments, the present invention is a method of treating aproliferative disorder, an immune disorder and a skin disorder in asubject in need thereof. The method comprises cutaneously administeringto the subject a therapeutically effective amount of a pharmaceuticalcomposition. The pharmaceutical composition comprises a pharmaceuticallyeffective amount of an active pharmaceutical ingredient (API) compoundrepresented by the following structural formula

at least one acidifying agent, and at least one pharmaceuticallyacceptable non-aqueous solvent. The values and preferred values of thevariables in structural formula (I) are as defined hereinabove.Particular and optional components and ingredients of pharmaceuticalcompositions useful in treating a proliferative disorder, an immunedisorder and a skin disorder according to the methods of the presentinvention have also been described above.

In some embodiments, the present invention is a method of treating aselected from a disorder selected from the group consisting of cutaneousT cell lymphoma, a skin cancer, a benign skin growth, acne, psoriasis,dermatitis, actinic keratosis, basal cell carcinoma, neurofibromatosis,a leukemia, a malignant melanoma, hair loss, and skin hyperpigmentation.In a particular embodiment, the disorder is selected from cutaneousT-cell lymphoma, neurofibromatosis, actinic keratosis, acne, basal cellcarcinoma, psoriasis, hair loss, skin pigmentation, and dermatitis.

Kits Comprising Pharmaceutical Compositions of the Present Invention

In some embodiments, the present invention is a kit, comprising a firstcontainer and a second container. The first container comprises anactive pharmaceutical ingredient (API) represented by the followingstructural formula

Values and preferred values of the variables in structural formula (I)are defined hereinabove. The second container comprises a vehicle base.The vehicle base includes at least one pharmaceutically acceptablenon-aqueous solvent and at least one acidifying agent. Particularpharmaceutically acceptable non-aqueous solvent and acidifying agentsare defined hereinabove.

In a particular embodiment, the first container comprises atherapeutically effective amount of the API.

In various embodiments, the kit can further include one or more of thefollowing components: at least one gelling agent, at least onehumectant, at least one emollient, at least one skin permeationenhancer, and at least one antioxidant. Suitable gelling agents,humectants, emollients, skin permeation enhancers and antioxidants aredefined hereinabove.

In some embodiments, the kit further includes a third containercomprising at least one gelling agent.

In some embodiments, the second container comprises ethanol, benzylalcohol, and citric acid. In other embodiments, the third containerfurther comprises propylene glycol, hexylene glycol, glycerin,diisopropyl adipate, oleyl alcohol, and butylated hydroxytoluene. In aparticular embodiment, the third container further comprises propyleneglycol, hexylene glycol, glycerin, diisopropyl adipate, oleyl alcohol,and butylated hydroxytoluene.

Any embodiment of the kit can further include instructions for the useof such kit. In some embodiments, the instructions include the steps ofadding the content of the first container to the second container. Inother embodiments, the instructions include the steps of blending thecontents of the first, the second and the third containers.

Referring to the FIGURE, one example of a manufacturing process that canbe employed to produce a pharmaceutical composition of the presentinvention is process 100. In step 102, solvents and emollients are addedto a vessel. In particular embodiments, solvents and emollients aremixed to uniformity. In step 104, one or more antioxidants are added tothe mixture. In a particular embodiment, butylated hydroxytoluene isadded to the mixture and dissolved. In step 106, an acidifying agent isadded to the vessel. In a particular embodiment, an anhydrous citricacid is added to the mixture. In step 108, an API is added to themixture. In step 110, a gelling agent is added to the vessel. In aparticular embodiment, Klucel MF PH is added to the mixture. In step112, the mixture is aliquoted into desirable containers. Optionally, instep 114, fill weight of each container is checked for accuracy.

EXEMPLIFICATION Example 1 Stability of Pharmaceutical Formulations ofthe Present Invention Materials and Methods

API used was the compound of structural formula (II). All excipientsused are listed in the United States Pharmacopeia-National Fromulary(USP/NF). Water and acetonitrile for HPLC analysis were HPLC grade. AnOrion 710A+ meter with a Ross Ultra electrode (Thermo Electron Corp.)was used to measure pH. The samples were diluted 1/10 prior to pHmeasurement to minimize junction potential errors in the measurement.Viscosity (at 20±1° C.) was measured with a Brookfield LVF viscometer at12 rpm using a #25 spindle and 13R sample holder. Since the samples werestored at different temperatures, they were allowed to equilibrate atthe viscosity measurement temperature for at least 12 hours. HPLCanalysis was performed using an HP 1050 system equipped with a variablewavelength UV/Vis detector. A gradient HPLC method is summarized inTable 1.

TABLE 1 Column ACE 5 micron C18 4.6 × 150 mm Mobile Phase A 10% ACN/90%,0.2% PCA Mobile Phase B 80% ACN/20%, 0.2% PCA Gradient 0.0 mm. 100% A20.0 mm. 100% B 20.1 mm. 100% A Run Time 28 min. Flow Rate 1.5 mL/min UVDetector 230 nm Injection Volume 20 μL Column Temperature 30° C. ACN:acetonitrile PCA: perchloric acid

Gel Preparation

One of ordinary skill in the art would understand that the process belowcan be modified, if necessary, in order to be scaled up.

The following laboratory-scale process has been used to manufacture gelsformulations of the pharmaceutical compositions of the presentinvention. (During this process, the vessel was open part of the time toallow insertion and operation of mixing blades attached to an overheadstirrer. To compensate for alcohol evaporation, a 2.0% overage of thissolvent was used.)

1. Solvents and emollients were added to a suitable vessel. A magneticstir bar was added, the vessel was sealed and placed on a stir plate.Mixing continued until the liquids are homogenous.

2. The seal was removed, BHT was added, and the seal was replaced. Thesolution was mixed until the antioxidant dissolved.

3. The seal was removed, the acid was added, and the seal was replaced.The solution was mixed until homogeneous. If citric acid was used, thepowder was added directly to the vessel. If phosphoric acid was used, itwas weighed into a small beaker then rinsed into the vessel with somereserved ethanol.

4. The seal was removed, compound (II) was added (for activeformulations only), and the seal was replaced. The solution was mixeduntil the active dissolved.

5. The seal was removed, the stir bar carefully removed, and a propellermixer blade attached to an overhead stirrer was inserted. The mixingspeed was adjusted to form a good vortex without excessive cavitation orsplashing.

6. The Klucel MF PH was slowly added into the vortex to disperse thepolymer. Mixing speed was adjusted as the formulation thickened tomaintain good mixing of the drug product without excessive aeration. Thevessel was partially covered to minimize evaporation.

7. Approximately 20-30 minutes after adding the polymer, thickeningoccurred to the point where the propeller blade was not sufficient tokeep all of the formulation mixing smoothly. The propeller blade wascarefully removed and the formulation was drained off the blade into thevessel (this was assisted with spatulation).

8. A suitable anchor blade attached to an overhead stirrer was insertedand mixing continued at a suitable speed. The vessel was partiallycovered to minimize evaporation.

9. Approximately 2 hours after adding the polymer, the gel washomogeneous with a slightly mottled appearance. To minimize evaporationwhile the polymer continued to swell, the anchor stirrer was carefullyremoved (allowing the gel to drain off into the vessel) and the vesselwas sealed.

10. The vessel was stored at room temperature protected from light for12-18 hours. After storage, the gel was remixed for an additional 15minutes. After final mixing was completed, the gel was filled into asuitable container.

Results

In neutral ethanol at room temperature, compound (II) degrades byapproximately 50% in 24 hours.

Six active batches were prepared at the 350 g scale. The citric acidlevel was reduced slightly so that the pH would move closer to 4.0.Their compositions and initial results are summarized in Tables 2A-2Band Tables 3A-3B, respectively.

TABLE 2A Compositions A, B, C and D (all values in % w/w, dehydratedethanol USP added to bring total to 100%) A B C D Compound (II) 1.0 1.01.0 1.0 Propylene glycol, USP 15.0 — 15.0 — Glycerin, USP 10.0 — 10.0 —diethylene glycol monoethyl — 25.0 — 25.0 ether, USP Hexylene glycol, NF12.0 12.0 12.0 12.0 Diisopropyl adipate 4.0 4.0 4.0 4.0 Oleyl alcohol,NF 5.0 5.0 5.0 5.0 BHT, NF 0.1 0.1 0.1 0.1 Citric acid (anhydrous), USP0.045 0.045 — — Phosphoric acid (85%), USP — — 0.015 0.015 Klucel MF PH2.0 2.0 2.0 2.0

TABLE 2B Compositions E and F (all values in % w/w) E F Compound (II)0.1 0.5 Propylene 15.0 15.0 glycol, USP Glycerin, USP 10.0 10.0Diethylene 0.0 0.0 glycol monoethyl ether, USP Hexylene glycol, 12.012.0 NF Diisopropyl 4.0 4.0 adipate Oleyl alcohol, 5.0 5.0 NF BHT, NF0.1 0.1 Citric acid 0.045 0.045 (anhydrous), USP Phosphoric acid 0 0(85%), USP Klucel MF PH 2.0 2.0 Dehydrated 51.755 51.355 alcohol(ethanol), USP

TABLE 3A Initial results for compositions in Table 2A A B C D Compound(II), 0.994 0.995 0.996 0.995 % w/w BHT assay, 0.099 0.099 0.102 0.101 %w/w pH 3.75 3.92 3.95 3.89 Viscosity, cP 18,800 20,000 17,600 21,200Appearance Conforms¹ Conforms¹ Conforms¹ Conforms¹ ¹Clear or translucentcolorless viscous liquid

TABLE 3B Initial results for compositions in Table 2B Property E FSHP-141 assay, % w/w 0.100 0.499 BHT assay, % w/w 0.099 0.099 pH 3.803.70 Viscosity, cP Not tested 24,532 Appearance Conforms¹ Conforms¹¹Clear to translucent colorless viscous liquid.

The results of a 6-month study of stability of formulations A, B and Care presented below in Table 4 through Table 8.

TABLE 4 Stability of Compound (II) gels after 2 weeks of storage at 40°C. A B C Compound (II), % initial 98.3 98.5 98.1 BHT, % initial 100.4100.1 99.7 pH 3.85 3.79 3.95 Viscosity, cP 16,800 17,600 16,000Appearance Conforms¹ Conforms¹ Conforms¹ ¹Clear or translucent colorlessviscous liquid

TABLE 5 Stability results for Compound (II) gels after 1 month ofstorage at 2-8 and 25° C. A B C Compound (II), % initial 1 month, 25° C.98.4 98.5 98.2 1 month, 2-8° C. 99.9 99.8 99.4 BHT assay, % initial 1month, 25° C. 99.8 100.3 99.8 1 month, 2-8° C. 99.7 99.9 99.6 pH 1month, 25° C. 3.92 3.80 3.90 1 month, 2-8° C. 3.84 3.97 3.99 Viscosity,cP 1 month, 25° C. 19,200 22,400 20,000 1 month, 2-8° C. 20,400 22,00021,600 Appearance 1 month, 25° C. Conforms¹ Conforms¹ Conforms¹ 1 month,2-8° C. Conforms¹ Conforms¹ Conforms¹ ¹Clear or translucent colorlessviscous liquid

TABLE 6 Stability of Compound (II) gels after 2 months of storage at 2-8and 25° C. A B C Compound (II), % initial 2 month, 25° C. 97.1 96.9 96.52 month 2-8° C. 99.1 98.8 98.6 BHT assay, % initial 2 month, 25° C. 99.599.7 99.5 2 month 2-8° C. 99.8 99.8 99.5 pH 2 month, 25° C. 3.85 3.914.02 2 month 2-8° C. 3.83 3.88 3.94 Viscosity, cP 2 month, 25° C. 20,40023,200 22,800 2 month 2-8° C. 20,800 21,200 23,200 Appearance 2 month,25° C. Conforms¹ Conforms¹ Conforms¹ 2 month 2-8° C. Conforms¹ Conforms¹Conforms¹ ¹Clear or translucent colorless viscous liquid

TABLE 7 Stability of Compound (II) gels after 3 months of storage at 2-8and 25° C. A B C Compound (II), % initial 3 months, 25° C. 96.4 96.195.6 3 months, 2-8° C. 98.6 98.3 97.5 BHT assay, % initial 3 months, 25°C. 99.7 99.7 99.6 3 months, 2-8° C. 99.5 99.4 99.9 pH 3 months, 25° C.3.95 3.81 3.96 3 months, 2-8° C. 3.82 3.84 4.05 Viscosity, cP 3 months,25° C. 24,000 24,400 24,000 3 months, 2-8° C. 22,800 23,200 24,800Appearance 3 months, 25° C. Conforms¹ Conforms¹ Conforms¹ 3 months, 2-8°C. Conforms¹ Conforms¹ Conforms¹ ¹Clear or translucent colorless viscousliquid

TABLE 8 Stability of Compound (II) gels after 6 months of storage at 2-8and 25° C. A B C Compound (II), % initial 6 month, 25° C. 95.0 94.5 93.46 month 2-8° C. 97.9 97.7 96.9 BHT assay, % initial 6 month, 25° C. 99.499.2 99.0 6 month 2-8° C. 99.3 99.4 99.3 pH 6 month, 25° C. 3.87 3.894.01 6 month 2-8° C. 3.85 3.94 3.95 Viscosity, cP 6 month, 25° C. 23,60023,600 25,200 6 month 2-8° C. 22,000 22,400 23,600 Appearance 6 month,25° C. Conforms¹ Conforms¹ Conforms¹ 6 month 2-8° C. Conforms¹ Conforms¹Conforms¹ ¹Clear or translucent colorless viscous liquid

The results of a 3-month study of stability of formulations E and F arepresented below in Table 9.

TABLE 9 Stability of formulations E and F after 3 months of storageProperty E F SHP-141 assay, % initial 3 month, 25° C. 100.1 99.0 3month, 2-8° C. 100.6 99.7 BHT assay, % initial 3 month, 25° C. 99.3103.3 3 month, 2-8° C. 100.6 103.8 pH 3 month, 25° C. 3.90 4.00 3 month,2-8° C. 3.90 4.00 Viscosity, cP 3 month, 25° C. 20,800 25,435 3 month,2-8° C. 21,550 25,288 Appearance 3 month, 25° C. Conforms¹ Conforms¹ 3month, 2-8° C. Conforms¹ Conforms¹ ¹Clear to translucent colorlessviscous liquid.

The results of a 6-month study of stability of formulation F arepresented below in Table 10.

TABLE 10 Stability of formulation F after 6 months of storage Property FSHP-141 assay, % initial 6 month, 25° C. 99.5 6 month, 2-8° C. 100.5 BHTassay, % initial 6 month, 25° C. 99.5 6 month, 2-8° C. 100.3 pH 6 month,25° C. 4.00 6 month, 2-8° C. 3.80 Viscosity, cP 6 month, 25° C. 25,448 6month, 2-8° C. 24,042 Appearance 6 month, 25° C. Conforms¹ 6 month, 2-8°C. Conforms¹ ¹Clear to translucent colorless viscous liquid.

The results of a 6-month study of stability of formulation E arepresented in Tables 11 and 12.

TABLE 11 Stability of formulation E after 6 months of storage at 5° C.Parameter E Assay 102.0 BHT 98.2 pH 3.9 Viscosity 21240 cP

TABLE 12 Stability of formulation E after 6 months of storage at 25° C.and 60% relative humidity Parameter E Assay 99.3 BHT 97.3 pH 3.9Viscosity 22480 cP

The results of a 12-month study of stability of formulation A and F arepresented in Tables 13 and 14.

TABLE 13 Stability of formulations A and F after 12 months of storage at5° C. Parameter F A Assay 102.5 100.9 BHT 99.2 99.2 pH 3.8 3.8 Viscosity29910 cP 23750 cP

TABLE 14 Stability of formulations A and F after 12 months of storage at25° C. and 60% relative humidity Parameter F A Assay 99.2 94.4 BHT 97.997.1 pH 3.9 3.8 Viscosity 27150 cP 25110 cP

Conclusions

In neutral ethanol at room temperature, compound (II) degrades byapproximately 50% in 24 hours.

It has now been discovered that compound (II) could be successfullyformulated in a acidified nonaqueous gel with acceptable cosmesis. Underthe typical temperature conditions in a refrigerated storage (2-8° C.),the formulation's saturation solubility for compound (II) wasapproximately 1.6% w/w. There was no significant change in appearance,pH, or BHT assay for 1% compound (II) gels after storage at 2-8° C. (3months), 25° C. (3 months), or 40° C. (2 weeks). The viscosity forvehicle and active gels showed a slight increase on storage at 2-8 or25° C., which is typical for nonaqueous Klucel gels. The slight increasein viscosity did an affect pourability or spreadability (assessed forvehicle gels, only). The compound (II) concentration decreased onaverage of by 1.9 or 4.0% after three months storage at 2-8 or 25° C.,respectively. Exposure to laboratory light for up to five days did notsignificantly affect the assay values for compound (II). All samplespassed identity and were clear, colorless, viscous gel solutions. Allassay, impurity, and BHT sample results are an average of threereplicate preparations, top, middle, and bottom, from single bottles.

Example 2 Formulation Kit Examples

Topical formulations of compound (II) were prepared from a kit thatcontains the following components: a vial of compound (II) powder, avial of solvent, and a container of gel concentrate. All three of thesecomponents are stable at controlled room temperature. At the time ofdispensing, the contents of the solvent vial were added to the compound(II) vial to dissolve the compound. After compound (II) had beensolubilized, the solution was added to the gel concentrate and thenmixed with a suitable implement (i.e., spatula) until homogeneous.

Three-compartment kit examples for two formulations are described belowin Table 15 and Table 16.

TABLE 15 Example Kit A Amount in Amount in Amount in Container 1,Container 2, Container 3, Compound % w/w % w/w % w/w Compound (II) 100 —— Ethanol (190 proof) — 84.95 — Propylene Glycol — 15.0 15.2 Citric Acid— 0.045 0.046 Hexylene Glycol — — 18.3 Glycerin — — 15.2 DiisopropylAdipate — — 6.10 Oleyl Alcohol — — 7.61 BHT — — 0.15 Klucel MF PH — —3.05 Ethanol (200 proof) — — 34.3

For 30 grams of gelled solution with a final compound (II) concentrationof 1%, the following weights of each component were used: 0.3 g ofcompound (II), 10.0 g of solvent, and 19.7 g of gel concentrate.

TABLE 16 Example Kit B Amount in Amount in Amount in Container 1,Container 2, Container 3, Compound % w/w % w/w % w/w Compound (II) 100 —— Ethanol (200 proof) — 74.95 34.9 Benzyl Alcohol — 25.0 — Citric Acid —0.045 0.046 Propylene Glycol — — 20.3 Hexylene Glycol — — 16.2 Glycerin— — 13.5 Diisopropyl Adipate — — 5.41 Oleyl Alcohol — — 6.76 BHT — —0.135 Klucel MF PH — — 2.70

For 30 grams of gelled solution with a final concentration of compound(II) of 1% using Formulation B, the following weights of each componentwere used: 0.3 g of compound (II), 7.5 g of solvent, and 22.2 g of gelconcentrate.

For both examples above, a homogeneous gelled formulation was obtainedafter less than 4 minutes of mixing. Compound (II) formulations preparedfrom a three compartment kit were stable for at least one month atcontrolled room temperature or in the refrigerator.

Example 3 Evaluation of the Percutaneous Absorption of Compound (II) InVitro Using the Franz Human Skin Finite Dose Model

In this example, solutions A and B, as described above, ewre used.Control solution (“Ctrl”) was a 1% by weight solution of compound (II)in DMSO.

Three test formulations containing compound (II)—A, B and Ctrl weretested on three replicate sections from two different ex vivo humantrunk skin donors, for the percutaneous absorption of compound (II), andfor the appearance of compound (III) depicted below, Methylparaben, and4-OH Benzoic acid over a 24 hour dose period. At preselected times afterdose application, the dermal receptor solution was removed in itsentirety, replaced with fresh receptor solution, and an aliquot savedfor subsequent analysis. In addition, the stratum corneum, epidermis anddermis were recovered and evaluated for drug content. The samples wereanalyzed for compound (II), compound of structural formula (III)

Methylparaben

and 4-OH Benzoic acid content by High Performance Liquid Chromatography(HPLC).

Materials and Methods

The in vitro Franz human skin finite dose model has proven to be avaluable tool for the study of percutaneous absorption and thedetermination of the pharmacokinetics of topically applied drugs. Themodel uses human ex vivo cadaver or surgical skin mounted in speciallydesigned diffusion cells that allow the skin to be maintained at atemperature and humidity that match typical in vivo conditions. A finitedose (e.g. 4-7 mg/cm²) of formulation is applied to the outer surface ofthe skin and drug absorption is measured by monitoring its rate ofappearance in the receptor solution bathing the inner surface of theskin. Data defining total absorption, rate of absorption, as well asskin content can be accurately determined in this model. The method hashistoric precedent for accurately predicting in vivo percutaneousabsorption kinetics.

Compound (II) degrades to less potent compounds, namely compound (III)and Methylparaben. Compound (III) may undergo further degradation tosuberic acid. Methylparaben may undergo degradation to 4-OH Benzoicacid. In this study compound (II), compound (III), Methylparaben and4-OH Benzoic acid were quantified in the collected samples.

Study Skin Preparation

Percutaneous absorption was measured using the in vitro Franz human skinfinite dose technique. Ex vivo, human trunk skin without obvious signsof skin disease, obtained within 24-48 hours of death, was used in thisstudy. It was dermatomed, prepared for cryopreservation, sealed in awater impermeable plastic bag, and stored at about −70° C. until the dayof the experiment. Prior to use it was thawed in about 37° C. water,then rinsed in water to remove any adherent blood or other material fromthe surface.

Skin from a single donor was cut into multiple smaller sections largeenough to fit on static 1.0 cm² Franz diffusion cells. The dermalchamber was filled to capacity with a reservoir solution ofphosphate-buffered isotonic saline (PBS), pH 7.4±0.1, and the epidermalcell (chimney) left open to ambient laboratory conditions.

All cells were mounted in a diffusion apparatus in which the dermalbathing solution was stirred magnetically at approximately 600 RPM andthe skin surface temperature maintained at 32.0° C.±1.0° C.

To assure the integrity of each skin section, its permeability totritiated water was determined before application of the test products.Following a brief (0.5-1 hour) equilibrium period, ³H₂O (NEN, Boston,Mass., sp. Act. apprx. 0.5 μCi/mL) was layered across the top of theskin so that the entire exposed surface was covered (approximately200-500 μL). After 5 minutes the ³H₂O aqueous layer was removed. At 30minutes the receptor solution was collected and analyzed for radioactivecontent by liquid scintillation counting. Skin specimens in whichabsorption of ³H₂O was less than 1.56 μL-equ/cm² were consideredacceptable.

Dosing and Sample Collection

Just prior to dosing, a pre-dose sample was taken and the reservoirsolution was replaced with a fresh solution of 0.1×PBS with 0.1% Volpoand 0.05% Citric acid (nominal pH 6.0). The chimney was removed from theFranz Cell to allow full access to epidermal the surface of the skin.All formulations were then applied to the skin sections using a positivedisplacement pipette set to deliver 5 μL formulation/cm². The dose wasevenly distributed onto the skin using a glass rod. The rod was retainedfor analysis to correct the applied dose. Five to ten minutes afterapplication the chimney portion of the Franz Cell was replaced. Atpreselected times after dosing, (6, 12, and 24 hours) the reservoirsolution was removed in its entirety, replaced with fresh reservoirsolution, and a predetermined volume aliquot saved for subsequentanalysis.

Spare cells were available which were not dosed but used to evaluate forthe appearance of substances diffusing out of the skin that mightinterfere with the analytic method.

After the last sample was collected, the surfaces were washed twice (0.5mL volume each) with Ethanol with 0.05% Citric Acid to collectun-absorbed formulation from the surface of the skin. Following thewash, the skin was tape stripped to remove the stratum corneum. The tapestrips were extracted overnight in neat acetonitrile. The skin was thenremoved from the chamber, split into epidermis and dermis. Each wasextracted overnight in a mixture of Ethanol and 0.05% citric acid overwet ice.

Analytical Methods

Samples were assayed using an HPLC/MS and/or HPLC/UV.

Results

Results are presented in Table 17 and Table 18, below. (In Tables 17 and18, an entry of “0” means that the parameter was below lower limit ofquantification.)

TABLE 17 Total Absorption and Mass Balance Results Across Skin Donors:Percutaneous Absorption and Penetration of Compound (II) Through ex vivoHuman Trunk Skin Over 24 Hours (Mean ± SE as Percent of Applied Dose andTotal Mass (μg/cm²).) 1% Compound 1% Compound 1% Compound (II) Gel (II)Gel (II) Solution Formulation Formulation (DMSO) A of Table 3 B of Table3 (Ctrl) Total Absorption 0.284 ± 0.028 0 0.296 ± 0.098 (μg/cm₂) Dermis(μg/cm₂) 0 0 0 Epidermis (μg/cm₂) 0.259 ± 0.074 0.151 ± 0.004 0.429 ±0.035 Stratum Corneum 0.048 ± 0.022 0.039 ± 0.039 0.016 ± 0.016 (μg/cm₂)Surface Wash 22.12 ± 3.938 31.94 ± 0.313 10.01 ± 0.189 (μg/cm₂) TotalAbsorption (%) 0.594 ± 0.060 0 0.600 ± 0.199 Dermis (%) 0 0 0 Epidermis(%) 0.542 ± 0.158 0.325 ± 0.004 0.870 ± 0.072 Stratum Corneum (%) 0.101± 0.045 0.087 ± 0.087 0.032 ± 0.032 Surface Wash (%) 46.28 ± 8.538 68.56± 1.957 20.29 ± 0.340 Total Recovery (%) 47.52 ± 8.592 68.97 ± 2.04821.79 ± 0.498

TABLE 18 Compound (II) Total Absorption Results Across DonorsPercutaneous: Absorption of Compound (II) Through ex vivo Human TrunkSkin Over 24 Hours From a Single Application. (Mean ± SE as Total Mass(μg/cm²) and Percent of Applied Dose.) Total Absorption Total AbsorptionTest Article (μg/cm2) (%) 1% Compound (II) Gel, 0.284 ± 0.028 0.594 ±0.060 Formulation A of Table 3 1% Compound (II) Gel, 0 0 Formulation Bof Table 3 1% Compound (II) Solution 0.296 ± 0.098 0.600 ± 0.199 (DMSO),Ctrl

Conclusion

Using the in vitro finite dose model, the data demonstrates thatcompound (II) penetrates into and through ex vivo human skin, fromformulations A and Ctrl, but not from formulation B (see Table 2 for theformulations A and B).

The penetration profile for compound (II) suggests that virtually allthe percutaneous absorption through the skin occurred within 5 hours ofdose application from formulations A and Ctrl, with no compound (II)being seen in the reservoir solution from formulation B. The majority ofthe detectable compound (II) was found in the surface wash (about20%-69%) followed by the epidermis (about 0.3%-0.9%).

The data further indicate that all of the degradation products to bequantified (compound (III), Methylparaben (structural formula (IV)), and4-OH Benzoic acid) were observed to be present in the majority of thesamples.

Further, degree of degradation was observed to be formulation dependentwith more degradation being seen in the samples dosed with formulationCtrl, followed by formulation A and formulation B.

Overall mass balance (based on the amount of compound (II) dose) as thesum across all measured compounds demonstrated about 71% recovery fromthe skin sections dosed with formulation B, about 55% from the skinsections dosed with formulation A, and about 51% from the skin sectionsdosed with formulation Ctrl. The unaccountable dose may represent otherdegradation products which were not quantified, or binding of selectedcompounds to the tissue or chamber system surfaces.

While this invention has been particularly shown and described withreferences to example embodiments thereof, it will be understood bythose skilled in the art that various changes in form and details may bemade therein without departing from the scope of the inventionencompassed by the appended claims.

1. A method of treating basal cell carcinoma in a subject in needthereof, said method comprising cutaneously administering to the subjecta therapeutically effective amount of a pharmaceutical compositioncomprising an active pharmaceutical ingredient (API) compoundrepresented by the following structural formula

at least one acidifying agent comprises citric acid; and a vehicle basecomprising at least one pharmaceutically acceptable non-aqueous solvent,wherein the pharmaceutical composition has a measured pH of from about 3to about 5; the API is present in an amount of about 0.5 to 5.0% (w/w);and the at least one non-aqueous solvent comprises ethanol.
 2. Themethod of claim 1 wherein the pharmaceutical composition furthercomprises at least one humectant.
 3. The method of claim 2, wherein theat least one humectant is selected from the group consisting of hexyleneglycol, glycerin, propylene glycol, sorbitol, lactic acid, sodiumlactate, mannitol, butylene glycol, panthenol, hyaluronic acid, urea,chitosan, polyols, methyl gluceth-10, methyl gluceth-20, andpolyethylene glycols.
 4. The method of claim 2, wherein the at least onehumectant is selected from glycerin and hexylene glycol.
 5. The methodof claim 1 wherein the pharmaceutical composition further comprises atleast one emollient.
 6. The method of claim 5 wherein the at least oneemollient of the pharmaceutical composition is selected from the groupconsisting of diisopropyl adipate, isopropyl myristate, isopropylpalmitate, cetearyl octonoate, isopropyl isostearate, myristyl lactate,octyldodecanol, oleyl alcohol, a mineral oil, petrolatum, a vegetableoil, PPG-15 stearyl ether, PEG-4 dilaurate, lecithin, lanolin, lanolinalcohol, polyoxyl 75 lanolin, cholesterol, cetyl esters wax, cetostearylalcohol, glyceryl monostearate, triglycerides of capric and caprylicacids, dimethicone, and cyclomethicone.
 7. The method of claim 5 whereinthe at least one emollient of the pharmaceutical composition is selectedfrom diisopropyl adipate and oleyl alcohol.
 8. The method of claim 1wherein the pharmaceutical composition further comprises at least onehumectant and at least one emollient.
 9. The method of claim 8 whereinthe at least one humectant of the pharmaceutical composition is selectedfrom glycerin and hexylene glycol and wherein the at least one emollientis selected from diisopropyl adipate and oleyl alcohol.
 10. The methodof claim 1 wherein the pharmaceutical composition further comprises atleast one skin permeation enhancer.
 11. The method of claim 10 whereinthe at least one permeation enhancer of the pharmaceutical compositionis selected from one or more of oleyl alcohol, proypylene glycol, andethanol.
 12. The method of claim 1 wherein the pharmaceuticalcomposition further comprises at least one gelling agent.
 13. The methodof claim 12, wherein the at least one gelling agent of thepharmaceutical composition is a hydroxypropylcellulose.
 14. The methodof claim 1 wherein the pharmaceutical composition further comprises atleast one antioxidant.
 15. The method of claim 14, wherein theantioxidant of the pharmaceutical composition is selected from the groupconsisting of alpha tocopherol, beta tocopherol, delta tocopherol, gammatocopherol, tocopherols, ascorbic acid, ascorbyl palmitate, butylatedhydroxyanisol (BHA), butylated hydroxytoluene (BHT), fumaric acid, malicacid, methionine, propyl gallate, sodium ascorbate, sodiummetabisulfate, sodium thiosulfate, sodium bisulfate.
 16. The method ofclaim 14, wherein the antioxidant is the butylated hydroxytoluene (BHT).17. The method of claim 1 wherein the pharmaceutical compositioncomprises at least 1.0% by weight of the API.
 18. The method of claim 1,wherein the ethanol is present in an amount of about 50% (w/w); andwherein the measured pH of the pharmaceutical composition is from about3 to about
 4. 19. The method of claim 17, wherein the ethanol is presentin an amount of about 50% (w/w); and wherein the measured pH of thepharmaceutical composition is from about 3 to about 4.